CD4+CD25+Foxp3+调节性T细胞在CIK细胞诱导中的表达及其功能

目的探讨肿瘤患者自体细胞因子诱导的杀伤细胞(CIK)诱导过程中CD4~+CD25~+Foxp3~+调节性T细胞的变化及其功能。方法应用血细胞分离机采集22例肿瘤患者外周血单个核细胞(PBMC),诱导培养CIK细胞,用流式细胞仪动态监测其CD4~+CD25~+Foxp3~+表型,并分析Tregs细胞负性调控分子转化生长因子-β1(TGF-β1)、细胞毒性T淋巴细胞相关抗原4(CTLA-4)和IL-10的表达水平,采用细胞增殖抑制试验测定Tregs细胞免疫学功能。结果诱导的CIK细胞中存在CD4~+CD25~+Foxp3~+Tregs,其表达量分别为第1天(0.30±0.15)%、第3天(4.48±1.72)%、第5天(3.83±2.12)%、第9天(2.37±1.17)%、第11天(1.65±0.99)%、第14天(1.04±0.76)%。诱导第14天时的Tregs细胞免疫调控负性分子TGF-β1的表达水平为(97.2±2.1)%、CTLA-4为(96.2±3.5)%、IL-10为(4.2±2.3)%。细胞增殖抑制试验中空白对照组、条件对照组以及实验组的增殖细胞表达量分别为(8.55±2.38)%、(42.66±7.32)%、(57.04±7.49)%。结论 CD4~+CD25~+Foxp3~+Tregs细胞可作为潜在的CIK细胞质量评价指标。     

CD4^＋CD25^＋调节性T细胞与自身免疫病

目的:CD4 CD25 调节性T细胞是一群具有免疫调节或免疫抑制功能的细胞。越来越多的实验证明,CD4 CD25 调节性T细胞在维持外周免疫耐受中起重要作用,这种T细胞的数量减少或功能缺失可导致自身免疫性疾病的发生。本文就CD4 CD25 调节性T细胞及其在自身免疫性疾病中作用的研究进展做一综述。资料来源:应用计算机检索CNKI、Medline、EMCC数据库和手工检索2006-2007年的相关文献。检索词为"CD4 CD25 T调节性细胞,自身免疫病,免疫耐受,CD4 CD25 regulatory T cell,Treg,autoimmune disease,immune tolerance"。资料选择:检索范围包括临床研究(不限研究对象的年龄、性别、种族)和基础研究,不限体内和体外研究。资料提炼:共收集到相关文献675篇,选择其中33篇英文文献进行重点阅读和分析。资料综合:CD4 CD25 调节性T细胞具有免疫抑制功能,在机体的免疫调节中发挥重要作用。与其免疫调节功能相关的杀伤性T细胞淋巴细胞相关抗原4、CD45RO、糖皮质激素诱导的肿瘤坏死因子受体、淋巴细胞的无能相关基因等细胞表面分子和白细胞介素2、白细胞介素10、白细胞介素4、转化生长因子β等细胞因子的研究不断深入。此外,CD4 CD25 调节性T细胞功能的发挥还与FOXP3的表达密切相关。CD4 CD25 调节性T细胞数量的减少、抑制功能的受损和(或)细胞表面分子表达的缺陷可能导致1型糖尿病、多发性硬化和炎症性肠病等多种自身免疫病的发生。结论:CD4 CD25 调节性T细胞主要通过细胞接触依赖机制和抑制性细胞因子依赖机制发挥免疫抑制效应。其数量的减少、功能的受损和(或)表面分子表达的缺陷与自身免疫病的发生发展密切相关。     

Antigen-dependent proliferation of CD4+ CD25+ regulatory T cells in vivo

The failure of CD25+ regulatory T cells (Tregs) to proliferate after T cell receptor (TCR) stimulation in vitro has lead to their classification as naturally anergic. Here we use Tregs expressing a transgenic TCR to show that despite anergy in vitro, Tregs proliferate in response to immunization in vivo. Tregs also proliferate and accumulate locally in response to transgenically expressed tissue antigen whereas their CD25- counterparts are depleted at such sites. Collectively, these data suggest that the anergic state that characterizes CD25+ Tregs in vitro may not accurately reflect their responsiveness in vivo. These observations support a model in which Treg population dynamics are shaped by the local antigenic environment.     

CD4+ FoxP3+ regulatory T cells confer infectious tolerance in a TGF-beta-dependent manner

CD4⁺FoxP3⁺ regulatory T (T reg) cells comprise a separate lineage of T cells that are essential for maintaining immunological tolerance to self. The molecular mechanism(s) by which T reg cells mediate their suppressive effects remains poorly understood. One molecule that has been extensively studied in T reg cell suppression is transforming growth factor (TGF)-β, but its importance remains controversial. We found that TGF-β complexed to latency-associated peptide (LAP) is expressed on the cell surface of activated but not resting T reg cells. T reg cell LAP–TGF-β plays an important role in the suppression of the proliferation of activated T cells, but it is not required for the suppression of naive T cell activation. More importantly, T reg cell–derived TGF-β could generate de novo CD4⁺FoxP3⁺ T cells in vitro from naive precursors in a cell contact–dependent, antigen-presenting cell–independent and α_V integrin–independent manner. The newly induced CD4⁺FoxP3⁺ T cells are suppressive both in vitro and in vivo. Transfer of activated antigen-specific T reg cells with naive antigen-specific responder T cells to normal recipients, followed by immunization, also results in induction of FoxP3 expression in the responder cells. T reg cell–mediated generation of functional CD4⁺FoxP3⁺ cells via this TGF-β–dependent pathway may represent a major mechanism as to how T reg cells maintain tolerance and expand their suppressive abilities.     

Cell contact-dependent immunosuppression by CD4(+)CD25(+) regulatory T cells is mediated by cell surface-bound transforming growth factor beta

CD4(+)CD25(+) T cells have been identified as a population of immunoregulatory T cells, which mediate suppression of CD4(+)CD25(-) T cells by cell-cell contact and not secretion of suppressor cytokines. In this study, we demonstrated that CD4(+)CD25(+) T cells do produce high levels of transforming growth factor (TGF)-beta1 and interleukin (IL)-10 compared with CD4(+)CD25(-) T cells when stimulated by plate-bound anti-CD3 and soluble anti-CD28 and/or IL-2, and secretion of TGF-beta1 (but not other cytokines), is further enhanced by costimulation via cytotoxic T lymphocyte-associated antigen (CTLA)-4. As in prior studies, we found that CD4(+)CD25(+) T cells suppress proliferation of CD4(+)CD25(-) T cells; however, we observed here that such suppression is abolished by the presence of anti-TGF-beta. In addition, we found that CD4(+)CD25(+) T cells suppress B cell immunoglobulin production and that anti-TGF-beta again abolishes such suppression. Finally, we found that stimulated CD4(+)CD25(+) T cells but not CD4(+)CD25(-) T cells express high and persistent levels of TGF-beta1 on the cell surface. This, plus the fact that we could find no evidence that a soluble factor mediates suppression, strongly suggests that CD4(+)CD25(+) T cells exert immunosuppression by a cell-cell interaction involving cell surface TGF-beta1.     

Human CD25+CD4+ T suppressor cell clones produce transforming growth factor beta,but not interleukin 10, and are distinct from type 1 T regulatory cells

T regulatory (Tr) cells are essential for the induction of peripheral tolerance. Several types of Tr cells exist, including CD4(+) T cells which express CD25 constitutively and suppress immune responses via direct cell-to-cell interactions, and type 1 T regulatory (Tr1) cells, which function via secretion of interleukin (IL)-10 and transforming growth factor (TGF)-beta. The relationship between CD25(+)CD4(+) T cells and Tr1 cells remains unclear. Here, we demonstrate at the clonal level that Tr1 and CD25(+)CD4(+) T cells are two distinct subsets of regulatory cells with different cytokine production profiles. Furthermore, CD25(-)CD4(+) T cells can be rendered anergic by IL-10 and differentiated into Tr1 cells in the absence of CD25(+)CD4(+) T cells. Cloned human CD25(+)CD4(+) T cell populations are heterogeneous and only a subset of clones continues to express high levels of CD25 and is suppressive. The intensity of CD25, cytotoxic T lymphocyte antigen (CTLA)-4, and glucocorticoid-induced tumor necrosis factor (TNF) receptor expression correlates with the suppressive capacity of the T cell clones. None of the CD25(+)CD4(+) T cell clones with suppressive function produce IL-10, but all produce TGF-beta. Suppression mediated by CD25(+)CD4(+) T cell clones is partially dependent on TGF-beta, but not on constitutive high expression of CD25. Together these data indicate that naturally occurring human CD25(+)CD4(+) T cells are distinct from IL-10-producing Tr1 cells.     

Tumor cells convert immature myeloid dendritic cells into TGF-beta-secreting cells inducing CD4+CD25+ regulatory T cell proliferation

The mechanisms through which regulatory T cells accumulate in lymphoid organs of tumor-bearing hosts remain elusive. Our experiments indicate that the accumulation of CD4+CD25+ regulatory T cells (T reg cells) expressing FoxP3 and exhibiting immunosuppressive function originates from the proliferation of naturally occurring CD25+ T cells and requires signaling through transforming growth factor (TGF)-beta receptor II. During tumor progression, a subset of dendritic cells (DCs) exhibiting a myeloid immature phenotype is recruited to draining lymph nodes. This DC subset selectively promotes the proliferation of T reg cells in a TGF-beta-dependent manner in mice and rats. Tumor cells are necessary and sufficient to convert DCs into regulatory cells that secrete bioactive TGF-beta and stimulate T reg cell proliferation. In conclusion, tumor expansion can stimulate T reg cells via a specific DC subset.     

脓毒症患儿CD4+CD25+调节性T细胞的变化

目的 观察不同免疫状态下脓毒症婴幼儿外周血CD4+CD25+Foxp3high调节性T细胞(Treg细胞)及相关分子的变化,探讨婴幼儿脓毒症免疫功能紊乱的可能机制.方法 分别收集2007年5月至2007年11月深圳市儿童医院重症监护室收治的婴幼儿脓毒症36例血液标本,另选16例健康同龄儿童作为正常对照进行前瞻性研究;排除既往患有自身免疫性疾病、免疫缺陷病、遗传代谢病及肿瘤的患儿,排除近6个月曾使用影响免疫功能的药物.本研究获得深圳市儿童医院伦理委员会的同意.以外周血CD14+单核细胞HLA-DR表达>30%或<30%为阈值,将患儿分为免疫激活组(DR-H组)和免疫抑制组(DR-L组),用流式细胞术检测CD14+单核细胞HLA-DR表达率,CD4+CD25+Foxp3highTreg细胞比例;实时荧光定量PCR(Real time-PCR)检测CD4+T细胞Foxp3、CTLA-4、GITR、IL-10mRNA表达.统计方法采用单因素方差分析,P<0.05为差异具有统计学意义.结果 急性期DR-L组CD4+CD25+Foxp3highTreg细胞比例明显高于对照组及DR-H组(P<0.05).DR-L组Foxp3、CTLA-4、IL-10等相关分子基因表达高于对照组及DR-H组(P<0.05),DR-L组GITR基因表达高于DR-H组.结论 CD4+CD25+Foxp3highTreg细胞数量异常增加可能与婴幼儿脓毒症免疫抑制状态有关.     

同种异基因抗原特异性调节性T细胞的体外扩增

目的建立用人B淋巴细胞体外获得大量的同种异基因抗原特异性调节性T(Treg)细胞的方法,以此检测扩增细胞的表型及其免疫无能性和免疫抑制性。方法第1轮扩增将免疫磁珠分选的人CD4+CD25+T细胞和人B淋巴细胞按1∶4体外混合培养,并加入外源白介素-2(IL-2)和抗-CD28;将第1轮扩增得到的Treg细胞用抗-CD3/CD28包被的免疫磁珠和IL-2刺激,做第2轮扩增以获得更多数量的抗原特异性Treg细胞,分为添加和或未添加免疫抑制剂雷帕霉素(RAPA)2组(n=3)。结果经过2轮的扩增后,在第2轮扩增中未添加RAPA组扩增1×103倍,纯度80%;添加RAPA组扩增0.8×103倍,纯度90%。添加RAPA组得到的Treg细胞的Foxp3、CT-LA4、CD39表达水平高于未添加RAPA组,但是HLA-DR变化不大;未添加RAPA组扩增得到的Treg细胞分泌低水平的IL-2、IL-17、IL-4和IFN-γ,而添加RAPA组得到的Treg细胞几乎不分泌上述各种细胞因子,前者表现出部分免疫反应无能性,后者表现出完全的免疫反应无能性,两者都表现出免疫抑制性功能特征。人B淋巴细胞扩增得到的抗原特异性Treg细胞能够极大地抑制同源抗原引起的免疫反应,而对多克隆刺激的免疫反应抑制能力较弱。结论用人B细胞体外扩增抗原特异性Treg细胞,再通过抗-CD3/CD28包被的免疫磁珠进一步刺激可以体外获得大量的抗原特异性的Treg细胞,加入RAPA后可有效地提高Treg细胞的纯度和免疫抑制力且呈现抗原特异性。     

Regulatory CD4+CD25+ T cells restrict memory CD8+ T cell responses

CD4+ T cell help is important for the generation of CD8+ T cell responses. We used depleting anti-CD4 mAb to analyze the role of CD4+ T cells for memory CD8+ T cell responses after secondary infection of mice with the intracellular bacterium Listeria monocytogenes, or after boost immunization by specific peptide or DNA vaccination. Surprisingly, anti-CD4 mAb treatment during secondary CD8+ T cell responses markedly enlarged the population size of antigen-specific CD8+ T cells. After boost immunization with peptide or DNA, this effect was particularly profound, and antigen-specific CD8+ T cell populations were enlarged at least 10-fold. In terms of cytokine production and cytotoxicity, the enlarged CD8+ T cell population consisted of functional effector T cells. In depletion and transfer experiments, the suppressive function could be ascribed to CD4+CD25+ T cells. Our results demonstrate that CD4+ T cells control the CD8+ T cell response in two directions. Initially, they promote the generation of a CD8+ T cell responses and later they restrain the strength of the CD8+ T cell memory response. Down-modulation of CD8+ T cell responses during infection could prevent harmful consequences after eradication of the pathogen.     

类风湿性关节炎患者外周血CD4^＋CD25^＋调节性T细胞的表达及意义

CD4 CD25 调节性T细胞是一种除主要表达CD25分子外,还表达CTLA-4和G ITR(糖皮质激素诱导的TNF受体)等的CD4 T细胞亚群,它不但是参与对自身抗原外周耐受的主要T细胞群,而且还作为对外来抗原应答的调节性T细胞,对于维持外周免疫耐受有重要意义[1,2]。已有研究表明,自身免疫病的易     

CD4~+CD25~+调节性T细胞的一般特性及其在银屑病发病中的作用

CD4+CD25+调节性T细胞是一组具有免疫调节作用的T细胞群,对于维持机体免疫耐受和免疫应答稳态具有非常重要的作用。越来越多的研究表明,银屑病是多基因遗传背景下T细胞介导的免疫性疾病,CD4+CD25+调节性T细胞的免疫反应及其分泌的细胞因子在银屑病的发病中有着重要作用。本文主要就CD4+CD25+调节性T细胞的一般特性及其在银屑病发病机制中作用的研究进展作一综述,帮助我们更深入地了解银屑病的发病机制并为今后临床诊断和治疗提供依据。     

CD4^＋CD25^＋调节性T细胞天冬氨酸特异性半胱氨酸蛋白酶-3／9活性改变及其与凋亡的关系

目的 探讨脓毒症时CD4+CD25+调节性T细胞(Treg)天冬氨酸特异性半胱氨酸蛋白酶-3/9(caspase-3/9)活性变化及其与激活诱导细胞凋亡的关系.方法 体外培养免疫磁珠法分选大鼠脾脏CD4+CD25+Treg,随机分为对照组、抗CD3/CD28组、抗CD3/CD28+脂多糖(LPS)组,培养3 d后采用流式细胞术检测Treg凋亡率,荧光比色法检测caspase-3/9活性.同时采用盲肠结穿孔术(CLP)复制大鼠脓毒症模型,将动物随机分为正常对照组、假手术组、CLP组,于第3日分离Treg后检测Treg凋亡率和caspase-3/9活性.结果 体外实验显示,抗CD3/CD28组和抗CD3/CD28+LPS组CD4+CD25+Treg凋亡率均显著高于对照组(P均<0.01),而抗CD3/CD28+LPS组Treg凋亡率明显低于抗CD3/CD28组(P<0.05).抗CD3/CD28组caspase-3/9活性均显著高于对照组(P均<0.01);抗CD3/CD28+LPS组caspase-3/9活性则明显低于抗CD3/CD28组(P均<0.01).体内实验显示,CLP组CD4+CD25+Treg凋亡率和caspase-3活性均显著低于正常对照组和假手术组(P均<0.01),但caspase-9活性改变不明显(P均>0.05).结论 脓毒症病理过程中CD4+CD25+Treg凋亡率明显下降,caspase-3激活是诱发Treg凋亡的重要途径之一.     

Direct expansion of functional CD25+ CD4+ regulatory T cells by antigen-processing dendritic cells

An important pathway for immune tolerance is provided by thymic-derived CD25+ CD4+ T cells that suppress other CD25- autoimmune disease-inducing T cells. The antigen-presenting cell (APC) requirements for the control of CD25+ CD4+ suppressor T cells remain to be identified, hampering their study in experimental and clinical situations. CD25+ CD4+ T cells are classically anergic, unable to proliferate in response to mitogenic antibodies to the T cell receptor complex. We now find that CD25+ CD4+ T cells can proliferate in the absence of added cytokines in culture and in vivo when stimulated by antigen-loaded dendritic cells (DCs), especially mature DCs. With high doses of DCs in culture, CD25+ CD4+ and CD25- CD4+ populations initially proliferate to a comparable extent. With current methods, one third of the antigen-reactive T cell receptor transgenic T cells enter into cycle for an average of three divisions in 3 d. The expansion of CD25+ CD4+ T cells stops by day 5, in the absence or presence of exogenous interleukin (IL)-2, whereas CD25- CD4+ T cells continue to grow. CD25+ CD4+ T cell growth requires DC-T cell contact and is partially dependent upon the production of small amounts of IL-2 by the T cells and B7 costimulation by the DCs. After antigen-specific expansion, the CD25+ CD4+ T cells retain their known surface features and actively suppress CD25- CD4+ T cell proliferation to splenic APCs. DCs also can expand CD25+ CD4+ T cells in the absence of specific antigen but in the presence of exogenous IL-2. In vivo, both steady state and mature antigen-processing DCs induce proliferation of adoptively transferred CD25+ CD4+ T cells. The capacity to expand CD25+ CD4+ T cells provides DCs with an additional mechanism to regulate autoimmunity and other immune responses.     

急性冠脉综合征患者CD4+CD25+调节性T细胞分析

动脉粥样硬化(AS)是涉及内皮细胞、巨噬细胞、平滑肌细胞和淋巴细胞间相互作用的多因素过程[1].斑块的增长影响器官血液灌流,导致脑血管事件和急性冠脉综合征(ACS)的发生.AS是一种慢性炎症性病变,受机体免疫反应的影响,抗原特异性效应T细胞(主要为CD4+T细胞)促进了斑块的产生和演变[2];调节性T细胞(Tregs)通过直接细胞接触机制和细胞因子调节机制对T细胞和B细胞产生调节作用,维持机体的免疫稳态;其中,CD4+CD25+Trges能通过对效应T细胞的抑制作用产生抗AS作用[3-4].为了解ACS患者外周血CD4+CD25+Trges数目变化和功能改变,我们检测了ACS患者外周血     

CD4+CD25+ regulatory T cells suppress allograft rejection mediated by memory CD8+ T cells via a CD30-dependent mechanism

CD4(+)CD25(+) regulatory T (Treg) cells suppress naive T cell responses, prevent autoimmunity, and delay allograft rejection. It is not known, however, whether Treg cells suppress allograft rejection mediated by memory T cells, as the latter mount faster and stronger immune responses than their naive counterparts. Here we show that antigen-induced, but not naive, Treg cells suppress allograft rejection mediated by memory CD8(+) T cells. Suppression was allospecific, as Treg cells induced by third-party antigens did not delay allograft rejection. In vivo and in vitro analyses revealed that the apoptosis of allospecific memory CD8(+) T cells is significantly increased in the presence of antigen-induced Treg cells, while their proliferation remains unaffected. Importantly, neither suppression of allograft rejection nor enhanced apoptosis of memory CD8(+) T cells was observed when Treg cells lacked CD30 or when CD30 ligand-CD30 interaction was blocked with anti-CD30 ligand Ab. This study therefore provides direct evidence that pathogenic memory T cells are amenable to suppression in an antigen-specific manner and identifies CD30 as a molecule that is critical for the regulation of memory T cell responses.     

调节性T细胞CD4~+CD25~+CD127~(low)在抗多耐药肺结核治疗中的意义

目的探讨调节性T细胞CD4+CD25+CD127low在西药联合中药复方"益肺通络方"抗多耐药结核患者中的免疫调节作用。方法选取多耐药结核病(MDR-TB)住院患者34例,随机分成标准化疗(西药)组12例作为对照组,中药复方联合西药组22例作为试验组,并签署知情同意书,按临床试验方案抗结核治疗,第0、3、6、9个月时分别采集2mL肝素抗凝外周静脉血,经CD4、CD25、CD127流式抗体标记后,Beckman流式细胞仪(FCM)计数各组CD4+CD25+CD127low百分比。结果随服药时间推移,对照组MDR-TB患者CD4+CD25+CD127low调节性T细胞百分比下降,差异无统计学意义(P0.05);试验组MDR-TB患者CD4+CD25+CD127low调节性T细胞百分比下降明显,差异有统计学意义(P=0.007);与对照组MDR-TB患者比较,试验组MDR-TB患者CD4+CD25+CD127low调节性T细胞百分比下降,差异有统计学意义(P=0.047)。结论经过9个月的抗结核治疗,试验组能有效降低MDR-TB患者CD4+CD25+CD127low调节性T细胞百分比,中药复方可能协同西药下调调节性T细胞的免疫抑制,恢复结核病患者机体免疫平衡状态。因此,中药复方联合西药抗多耐药结核患者治疗也许是一个有益的帮助。